Neutrophil activation by nanomaterials in vitro: comparing strengths and limitations of primary human cells with those of an immortalized (HL-60) cell line

ABSTRACT We previously reported that mitogenic activation of porcine peripheral blood mononuclear cells resulted in production of porcine endogenous retrovirus(es) (PERV[s]) capable of productively infecting human cells (C. Wilson et al., J. Virol. 72:3082–3087, 1998). We now extend that analysis to show that additional passage of isolated virus, named here PERV-NIH, through a human cell line yielded a viral population with a higher titer of infectious virus on human cells than the initial isolate. We show that in a single additional passage on a human cell line, the increase in infectivity for human cells is accounted for by selection against variants carrying pig-tropic envelope sequences (PERV-C) as well as by enrichment for replication-competent genomes. Sequence analysis of the envelope cDNA present in virions demonstrated that the envelope sequence of PERV-NIH is related to but distinct from previously reported PERV envelopes. The in vitro host range of PERV was studied in human primary cells and cell lines, as well as in cell lines from nonhuman primate and other species. This analysis reveals three patterns of susceptibility to infection among these host cells: (i) cells are resistant to infection in our assay; (ii) cells are infected by virus, as viral RNA is detected in the supernatant by reverse transcription-PCR, but the cells are not permissive to productive replication and spread; and (iii) cells are permissive to low-level productive replication. Certain cell lines were permissive for efficient productive infection and spread. These results may prove useful in designing appropriate animal models to assess the in vivo infectivity properties of PERV.

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SELECTION AND TESTING OF EXPERIMENTAL MODELS OF NORMAL AND MALIGNANT HUMAN CELLS IN VITRO AND EVALUATION OF THEIR SENSITIVITY RANGE TO THE NEUTRON/CAPTURE AND PHOTON-CAPTURE AGENTS AND PHOTOSENSITIZERS

Проблеми радіаційної медицини та радіобіології = Problems of Radiation Medicine and Radiobiology ◽

10.33145/2304-8336-2021-26-260-272 ◽

2021 ◽

Vol 26 ◽

pp. 260-272

Author(s):

O. Pochapinskyi ◽

◽

G. Lavrenchuk ◽

N. Atamaniuk ◽

A. Chernyshov ◽

...

Keyword(s):

Cell Line ◽

Neutron Capture ◽

Human Fibroblasts ◽

Cancer Cell Line ◽

Lung Cancer Cell Line ◽

Human Cells ◽

In Vitro Test ◽

Malignant Cells ◽

Test Systems

Objective: to investigate the structural and morpho-functional changes in test systems of malignant (A-549 cell line) and normal (fibroblasts of the 6th passage) human cells during incubation with gadolinium-containing photon-capture agent «Dotavist» and photosensitizer «Fotolon». Methods. The passaged (continuously interweaved) cell culture technique on normal human fibroblasts and malignant human cells; cytological, biophysical, statistical methods. Results. The cytotoxic properties of «Dotavist» gadolinium-containing photon-capturing agent and «Photolon» photosensitizer in a wide range of concentrations (5, 10, 25, 50, 100 and 200 μl/ml) were studied by the morphofunctional characteristics (growth kinetics, proliferative and mitotic activity, presence of atypical cells) in the in vitro test systems of malignant (non-small cell lung cancer cell line A-549) and normal (6th passage fibroblasts) human cells. It was found that the cytotoxic properties of «Dotavist» in test systems of malignant and normal cells are expressed under its administration in high concentrations (100 and 200 μl/ml). During incubation with «Photolon» photosensitizer the cytotoxic effect on malignant cells was determined at the lowest concentrations (5 and 10 μl/ml). Photosensitizer administration in the increasing concentrations has lead to genotoxic effects. Cytotoxic effect of photosensitizer on the normal human fibroblasts was evident in the 5-200 μl/ml concentration range. There was a moderate decrease in mitotic activity along with increasing concentration. Genotoxic properties of photosensitizer were evident at 25 μl/ml concentration and above. Conclusion. Study results of the effectiveness of neutron-capture and photon-capture technologies by the sensitivity assay in the in vitro test systems of human malignant cells (non-small cell lung cancer cell line A-549) and normal cells (transplantable human fibroblast culture, the 6th passage) to the gadolinium-containing photon-capture «Dotavist» agent and «Photolon» photosensitizer in different concentrations provide the basis for pre-clinical stage of evaluating the effectiveness of medications used in binary technologies. Key words: culture of human malignant cells, culture of human fibroblasts, neutron-capture agent, photon-capture agent, photosensitizer, proliferation, mitotic index.

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A CellAgeClock for expedited discovery of anti-ageing compounds

10.1101/803676 ◽

2019 ◽

Cited By ~ 2

Author(s):

Celia Lujan ◽

Eleanor J. Tyler ◽

Simone Ecker ◽

Amy P. Webster ◽

Eleanor R. Stead ◽

...

Keyword(s):

Cpg Methylation ◽

Human Cells ◽

Epigenetic Clock ◽

The Novel ◽

Drug Treatments ◽

Screening Platform ◽

Methylation Profiling ◽

Primary Human Cells

AbstractWe aim to improve anti-ageing drug discovery, currently achieved through laborious and lengthy longevity analysis. Recent studies demonstrated that the most accurate molecular method to measure human age is based on CpG methylation profiles, as exemplified by several epigenetics clocks that can accurately predict an individual’s age. Here, we developed CellAgeClock, a new epigenetic clock that measures subtle ageing changes in primary human cells in vitro. As such, it provides a unique tool to measure effects of relatively short pharmacological treatments on ageing. We validated the CellAgeClock against known longevity drugs such as rapamycin and trametinib. Moreover, we uncovered novel anti-ageing drugs, torin2 and Dactolisib (BEZ-235), demonstrating the value of our approach as a screening and discovery platform for anti-ageing strategies. The CellAgeClock outperforms other epigenetic clocks in measuring subtle ageing changes in primary human cells in culture. The tested drug treatments reduced senescence and other ageing markers, further consolidating our approach as a screening platform. Finally, we show that the novel anti-ageing drugs we uncovered in vitro, indeed increased longevity in vivo. Our method expands the scope of CpG methylation profiling from measuring human chronological and biological age from human samples in years, to accurately and rapidly detecting anti-ageing potential of drugs using human cells in vitro, providing a novel accelerated discovery platform to test sought after geroprotectors.

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Protective effects of blue light-blocking shades on phototoxicity in human ocular surface cells

BMJ Open Ophthalmology ◽

10.1136/bmjophth-2018-000217 ◽

2019 ◽

Vol 4 (1) ◽

pp. e000217 ◽

Cited By ~ 4

Author(s):

Yoshimi Niwano ◽

Atsuo Iwasawa ◽

Kazuo Tsubota ◽

Masahiko Ayaki ◽

Kazuno Negishi

Keyword(s):

Blue Light ◽

Ocular Surface ◽

Human Cells ◽

Light Irradiation ◽

Protective Effects ◽

Corneal Surface ◽

Surface Protection ◽

Viable Cells ◽

Primary Human Cells

ObjectiveBlue light hazards for retina and ocular surface have been repeatedly described and many protective methods are introduced for retina; however, no study has been conducted on ocular surface protection. The purpose of this in vitro study was to examine phototoxicity and shade protection after blue light irradiation in primary human cells of corneal surface origin.Methods and analysisPrimary human cells of corneal surface origin were obtained from eye bank eyes. After blue light irradiation (405 nm) of these cells for 3 min, and a further 24 hours’ incubation, surviving viable cells were assessed by the methyl thiazolyl tetrazolium assay. Simultaneously, cell viability was determined in wells covered by ultraviolet and blue light shades.ResultsUnder subconfluent conditions, viable cells decreased by around 50% after blue light irradiation, compared with control cells without irradiation. The blue light phototoxicity was not blocked by the control shade, but the ultraviolet-blocking and blue light-blocking shades protected the cells from phototoxicity, producing a 30%–40% reduction (ultraviolet) and 15%–30% reduction (blue light) in viable cells.ConclusionThese results indicate that blue light injures ocular surface cells and the cells are protected from damage by a shade. We recommend blue light protection to maintain ocular health, especially in high-risk populations, such as people with dry eye, contact lens users, the malnourished and the elderly.

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Assessment of Cell Line Models of Primary Human Cells by Raman Spectral Phenotyping

Biophysical Journal ◽

10.1016/j.bpj.2009.12.4289 ◽

2010 ◽

Vol 98 (8) ◽

pp. 1703-1711 ◽

Cited By ~ 63

Author(s):

Robin J. Swain ◽

Sarah J. Kemp ◽

Peter Goldstraw ◽

Teresa D. Tetley ◽

Molly M. Stevens

Keyword(s):

Cell Line ◽

Human Cells ◽

Raman Spectral ◽

Primary Human Cells

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An In Vitro and In Vivo Evidence That Downregulation of Leukemia Inhibitory Factor (LIF) Receptor (LIF-R) Decreases the Metastatic Potential of Human Rhabdomyosarcoma (RMS) Cells.

Blood ◽

10.1182/blood.v108.11.2561.2561 ◽

2006 ◽

Vol 108 (11) ◽

pp. 2561-2561

Author(s):

Marcin Wysoczynski ◽

Katarzyna Miekus ◽

Anna Marcinkowska ◽

Anna Janowska-Wieczorek ◽

Mariusz Z. Ratajczak

Keyword(s):

Skeletal Muscle ◽

Bone Marrow ◽

Cell Line ◽

Cell Lines ◽

Leukemia Inhibitory Factor ◽

Biological Significance ◽

Human Cells ◽

Inhibitory Factor

Abstract Rhabdomyosarcoma (RMS) and skeletal muscle-derived tumors frequently infiltrate bone marrow (BM). We have demonstrated that the stromal-derived factor (SDF)-1-CXCR4 receptor (Blood2002;100:2597) and hepatocyte growth factor (HGF)-c-Met receptor (Cancer Res. 2003;63:7926) play an important role in RMS metastasis to BM. Leukemia inhibitory factor (LIF) is a well known factor that plays an important role in skeletal muscle development/regeneration and similarly as SDF-1 and HGF is secreted by BM stroma. This prompted us to examine whether the LIF-LIF receptor (LIF-R) axis affects the biology/metastasis of RMS cells. We employed in our studies, human established RMS cell lines, as well as RMS samples isolated from patients and noticed that LIF-R was expressed not only on established human RMS cell lines (7/7) but more importantly, it was also detectable in patient samples (23/23). We also found that in RMS cells LIF stimulatesphosphorylation of MAPKp42/44, AKT and STAT3,chemotaxis and adhesion andincreases resistance to cytostatics (e.g., etoposide). These LIF-mediated effects were inhibited after downregulating the LIF-R by siRNA. To learn more on the biological significance of the LIF-LIF-R axis in vivo we employed two models. First, human RMS cells (RH-30) were exposed or not exposed to LIF-R siRNA and subsequently injected into SCID™-Beige immunodeficient mice. To estimate the number of RMS cells that seed to BM and liver in these animals, we isolated DNA and using real- time RT-PCR, amplified human a-satellite sequences and murine b-actin. The number of human cells present in murine organs was subsequently calculated from a standard curve derived from mixing varying numbers of human cells with a constant number of murine cells. We noticed that downregulation of LIF-R by siRNA significantly decreased the number of human RMS cells in murine BM and liver (x4 and x2 respectively). In a second model, the RH30 cell line was selected by repetitive chemotaxis for cells that are highly responsive to LIF (RH-30 L) and subsequently the cells from parental RH-30 cell line and RH-30 L cells were injected intramuscularly. Six weeks after tumour inoculation, we detected more metastasis in bone marrow and lungs in mice injected with RH-30L cells as compared to parental RH-30 clone (x6 and x3 respectively). In conclusion, we present evidence for the first time that the inhibition of LIF-LIF-R axis may decrease the invasive potential of human RMS both in vitro and in vivo. Hence, molecular targeting of LIF-LIF-R axis could possibly become a more effective new strategy to control the progression and metastasis of RMS.

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Influence of traditionally used Nepalese plants on wound healing and immunological properties using primary human cells in vitro

Journal of Ethnopharmacology ◽

10.1016/j.jep.2019.02.034 ◽

2019 ◽

Vol 235 ◽

pp. 415-423 ◽

Cited By ~ 3

Author(s):

Amy M. Zimmermann-Klemd ◽

Viktoria Konradi ◽

Carmen Steinborn ◽

Annekathrin Ücker ◽

Chiara Madlen Falanga ◽

...

Keyword(s):

Wound Healing ◽

Human Cells ◽

Immunological Properties ◽

Primary Human Cells

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Mitotic non-disjunction as a mechanism for in vitro aneuploidy induction by X-rays in primary human cells

Mutagenesis ◽

10.1093/mutage/11.4.307 ◽

1996 ◽

Vol 11 (4) ◽

pp. 307-313 ◽

Cited By ~ 36

Author(s):

M. Kirsch-Volders ◽

I. Tallon ◽

C. Tanzarella ◽

A. Sgura ◽

T. Hermine ◽

...

Keyword(s):

Human Cells ◽

X Rays ◽

Primary Human Cells

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Neutrophil trafficking on-a-chip: an in vitro, organotypic model for investigating neutrophil priming, extravasation, and migration with spatiotemporal control

Lab on a Chip ◽

10.1039/c9lc00562e ◽

2019 ◽

Vol 19 (21) ◽

pp. 3697-3705 ◽

Cited By ~ 1

Author(s):

Patrick H. McMinn ◽

Laurel E. Hind ◽

Anna Huttenlocher ◽

David J. Beebe

Keyword(s):

Human Cells ◽

Microfluidic Technology ◽

And Migration ◽

High Degree ◽

Spatiotemporal Control ◽

Neutrophil Priming ◽

Primary Human Cells

Her we report a new microfluidic technology designed to facilitate the study of neutrophil trafficking and priming using primary human cells with a high degree of spatiotemporal control.

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